Principal Investigator/Program Director (Last, First, Middle): Chaiken, Irwin M / Freire, Ernesto DESCRIPTION: The viral envelope glycoprotein gp120 is responsible for the initial events in HIV-1 infection. The binding of gp120 to the cell surface receptor CD4 induces a series of allosteric events that culminate with the fusion of the viral and cell membranes. The entire series of events defines different intervention points at which the process can be interrupted, therefore providing various opportunities for inhibitor development. These intervention points define two classes of inhibitors: compounds that competitively inhibit the binding of gp120 to its cellular partners and compounds that block allosteric signaling and downstream activation. The rational design and optimization of.inhibitors directed at any of these intervention points requires a precise knowledge of the structural energetics and conformational stability of gp120, its binding interactions, allosteric pathways and potential sites for inhibitor targeting. In addition, since the vast majority of HIV-1 infection occurs in Sub-Saharan Africa, where the main viral subtypes are C and A rather than the B subtype responsible for the infection in the United States and Western Europe, it is important that inhibitors are effective against those subtypes since they differ by as much as 30% in their genomes, including gp120. these are the main issues addressed in this project and can be summarized in the following specific aims: 1. Thermodynamic characterization of the structural stability and cooperative linkage between binding sites in gp120 by utilizing a combination of microcalorimetric and structure-based thermodynamic studies. 2. Development of thermodynamic guidelines for competitive inhibitors. How do we inhibit gp120 binding without triggering the allosteric activation cascade? 3. Development of thermodynamic guidelines for allosteric inhibitors. Identification of potential binding sites that can be targeted for blocking gp120 allosteric pathways. 4. Characterization of the allosteric inhibitor BMS-806 and analogs, the only gp120 inhibitor in clinical studies. Identification of the BMS-806 binding site. Identification of the critical functionalities in BMS- 806. Development of BMS-806-like inhibitors based upon different chemical scaffolds. 5. Development of thermodynamic guidelines for gp120 inhibitors that are effective against different HIV-1 subtypes and exhibit low susceptibility to potential drug resistant mutants. The studies involve a combination of experimental thermodynamic measurements (isothermal titration calorimetry and differential scanning calorimetry) and structure-based thermodynamic analysis.